Epidemiology of vancomycin-resistant enterococci in Canadian hospitals (CANWARD study, 2007 to 2013).

Of 1,927 Enterococcus species isolates collected across Canada from 2007 to 2013, 80 (4.2%) were identified as vancomycin-resistant enterococci (VRE). VRE infections during this time tripled in Canadian hospitals, from 1.8% to 6.0% (P = 0.03). All VRE were Enterococcus faecium, with 90% possessing vanA. The prevalence of vanB decreased from 37.5% in 2007 to 0% in 2013 (P < 0.05). The VRE were multidrug resistant, but 70.6%, 86.3%, and 100% were susceptible to doxycycline, linezolid, and daptomycin, respectively.

Distinguishable epidemics of multidrug-resistant Salmonella Typhimurium DT104 in different hosts.

The global epidemic of multidrug-resistant Salmonella Typhimurium DT104 provides an important example, both in terms of the agent and its resistance, of a widely disseminated zoonotic pathogen. Here, with an unprecedented national collection of isolates collected contemporaneously from humans and animals and including a sample of internationally derived isolates, we have used whole-genome sequencing to dissect the phylogenetic associations of the bacterium and its antimicrobial resistance genes through the course of an epidemic. Contrary to current tenets supporting a single homogeneous epidemic, we demonstrate that the bacterium and its resistance genes were largely maintained within animal and human populations separately and that there was limited transmission, in either direction. We also show considerable variation in the resistance profiles, in contrast to the largely stable bacterial core genome, which emphasizes the critical importance of integrated genotypic data sets in understanding the ecology of bacterial zoonoses and antimicrobial resistance.

Public health genomics and the new molecular epidemiology of bacterial pathogens.

Laboratory methods that can unambiguously fingerprint pathogenic microbes are needed to investigate the transmission of human infectious diseases from diverse sources, such as from the community, from the environment, within hospitals, or from contaminated food or water sources. Public health investigations currently rely on laboratory subtyping methods that ultimately provide only a fraction of the total genetic information of a pathogen, and although there is widespread success using existing subtyping methods, they do not always provide sufficient evidence to link disease cases together into outbreaks or to link these human cases to the culprit source. Alternatively, whole-genome sequencing of bacterial pathogens provides an unabridged examination of the genetic content of individual pathogen isolates, enabling public health laboratories to benefit from comparative analyses of total genetic content. In this context, whole-genome sequencing represents the ultimate epidemiological typing method - a universally applicable, highly detailed typing platform capable of providing the entire genetic blueprint of a pathogen and distinguishing strains to the single nucleotide level. These new genomic methods, if implemented within existing public health laboratory response programs, promise to revolutionize the ability of the laboratory to provide information and evidence on the evolution, transmission and virulence for bacterial pathogens - and this revolution is launching the new field of 'genomicepidemiology'.

Evaluation of a new chromogenic agar medium for detection of Shiga toxin-producing Escherichia coli (STEC) and relative prevalences of O157 and non-O157 STEC in Manitoba, Canada.

This study assesses the detection performance of CHROMagar STEC medium relative to a reference cytotoxin assay and describes the current relative prevalence of O157 and non-O157 Shiga toxin-producing Escherichia coli (STEC) serotypes within the province of Manitoba, Canada. Over a 10-month period, 205 nonfrozen routine stool submissions to Cadham Provincial Laboratory (CPL) were used to assess the performance of CHROMagar STEC. Of the 205 stools, 14 were identified as true positives by a cytotoxin assay, with resultant CHROMagar STEC sensitivity, specificity, and positive predictive and negative predictive values of 85.7%, 95.8%, 60.0%, and 98.9%, respectively. Using a separate panel of 111 STEC strains, CHROMagar STEC was shown to support the growth of 96 (86.5%) isolates. To assess relative prevalence, attempts were made to isolate by any means all STEC strains identified at CPL over a 17-month period. Of 49 isolates (representing 86.0% of all STEC infections detected), only 28.6% were O157 STEC strains. Of the 35 non-O157 STEC strains, 29 were subjected to further molecular analysis. In contrast to earlier results from our area, carriage of stx(2) appears to have increased. Overall, although CHROMagar STEC is not recommended as a primary screen, our results indicate that it is an effective supplemental medium for the isolation of probable STEC strains. Increased isolation of these serotypes is warranted to better understand their prevalence, clinical characteristics, and epidemiology and aid in the development or enhancement of food safety control programs targeting all STEC serotypes.

Is there a reservoir of sub-clinical lymphogranuloma venereum and non-LGV Chlamydia trachomatis infection in men who have sex with men?

SUMMARY: The aim of this study was to determine if a reservoir of sub-clinical LGV infection exists in men who have sex with men (MSM), as this finding might account for the recent rise in lymphogranuloma venereum (LGV) Chlamydia trachomatis infections among MSM in Canada. MSM without proctitis were enrolled between January and August 2006 in a cross-sectional study. Rectal, urine, serology and pharyngeal specimens were tested for specific C. trachomatis serovars. The median age of the 253 participants was 43 years; 53% were HIV+. We found no active cases of LGV infection; but 20 (8%) participants had positive serology. Thirteen participants (5%) had non-LGV C. trachomatis infections. Unprotected anopenetrative intercourse, rectal enema and drug use were associated with non-LGV C. trachomatis infection. Sub-clinical rectal non-LGV C. trachomatis infection was relatively common but LGV was not identified in our sample. Further studies of screening for non-LGV chlamydia infection in MSM are needed.

Cellular location and temperature-dependent assembly of IncHI1 plasmid R27-encoded TrhC-associated conjugative transfer protein complexes.

Conjugal transfer of IncHI plasmid DNA between Gram-negative bacteria is temperature sensitive, as mating is optimal between 22 degrees C and 30 degrees C but is inhibited at 37 degrees C. R27, isolated from Salmonella enterica serovar Typhi, is an IncHI1 plasmid of 180 kbp that has been sequenced completely. The gene encoding green fluorescent protein (GFP) was inserted into R27 in frame with trhC. TrhC is a mating pair formation (Mpf) protein that is essential for plasmid transfer and H-pilus production. Fluorescence microscopy allowed visualization of the TrhC-GFP fusion protein, and Escherichia coli cells were examined for the subcellular localization and temperature-dependent production of TrhC-GFP. At 27 degrees C, TrhC-GFP was found at the periphery of cells as discrete foci, indicating an association of TrhC within protein complexes in the bacterial cell membrane, whereas at 37 degrees C, little fluorescence was detected. These foci probably represent the intracellular position of protein complexes involved in conjugative transfer, as the formation of foci was dependent upon the presence of other Mpf proteins. During temperature shift experiments from 37 degrees C to 27 degrees C, a long lag period was required for generation of GFP foci. Conversely, during short shifts from 27 degrees C to 37 degrees C, the GFP foci remained stable. These results suggest that the expression of transfer genes in the Tra2 region of R27 is temperature dependent. Subcellular localization of TrhC was verified by cellular fractionation. Expression patterns of TrhC-GFP were confirmed with immunoblot analysis and reverse transcriptase-polymerase chain reaction (RT-PCR). These results allow us to propose mechanisms to explain the temperature-sensitive transfer of R27.

The complete DNA sequence and analysis of R27, a large IncHI plasmid from Salmonella typhi that is temperature sensitive for transfer.

Salmonella typhi, the causative agent of typhoid fever, annually infects 16 million people and kills 600 000 world wide. Plasmid-encoded multiple drug resistance in S. typhi is always encoded by plasmids of incompatibility group H (IncH). The complete DNA sequence of the large temperature-sensitive conjugative plasmid R27, the prototype for the IncHI1 family of plasmids, has been compiled and analyzed. This 180 kb plasmid contains 210 open reading frames (ORFs), of which 14 have been previously identified and 56 exhibit similarity to other plasmid and prokaryotic ORFs. A number of insertion elements were found, including the full Tn 10 transposon, which carries tetracycline resistance genes. Two transfer regions, Tra1 and Tra2, are present, which are separated by a minimum of 64 kb. Homologs of the DNA-binding proteins TlpA and H-NS that act as temperature-regulated repressors in other systems have been located in R27. Sequence analysis of transfer and replication regions supports a mosaic-like structure for R27. The genes responsible for conjugation and plasmid maintenance have been identified and mechanisms responsible for thermosensitive transfer are discussed.